The term cachexia embraces a complex metabolic syndrome characterized by loss of body weight that may develop as a consequence
of loss of muscle mass with or without loss of fat mass; bone mineral density may be affected as well [1].
Over the last 10 years, the Cachexia Conference has developed a
forum for researchers from the fields of cachexia and wasting
disorders. It is unique in several ways as it provides a
platform for both clinicians and basic researchers to meet and discuss
pathways and potential therapeutic targets as well as
recent evidence from clinical trials. The 6th Cachexia Conference was
held in Milan, Italy, from 8 to 10 December 2012 with
over 400 participants from more than 25 countries attending [2].

Cachexia remains underdiagnosed and
undertreated, even though it is prevalent among many patients presenting
with cancer,
chronic heart failure, chronic obstructive pulmonary
disease, human immunodeficiency virus infection or chronic kidney
disease.
Creating awareness for cachexia is one of the major aims
of the Cachexia Conference as well as the improvement of patients'
morbidity, mortality and quality of life. A more thorough
understanding of the underlying mechanisms and pathophysiology may
help in achieving these aims. The identification of such
mechanisms may help to identify therapeutic targets and potential
biomarkers that help to detect loss of tissue as early as
possible. Many excellent animal and clinical studies were presented
at the meeting in Milan, and the following overview seeks
to highlight some major research areas in the field of cachexia
and body wasting.

2 Muscle mitochondrial dysfunction

A number of elegant models were presented
in order to improve our understanding of pathways involved in the
wasting process.
Muscle wasting has received increasing research efforts
in recent years. Thus, one of the hot topics of the meeting was the
investigation of muscle proteolytic pathways. Slimani et
al. from Didier Attaix's group (Clermont University, Clermont-Ferrand,
France) used a rat model of immobilisation-induced muscle
atrophy to examine underlying pathomechanisms. They demonstrated
that proteolysis via the ubiquitin proteasome system
(UPS) and mitochondrium-associated apoptosis are involved in muscle
remodelling
during early recovery in immobilised muscle [3].
In addition, these authors demonstrated that the muscle-specific E3
ubiquitin ligase muscle RING-finger protein 1 (MuRF1)
is up-regulated during catabolic conditions and that it
is involved in the polyubiquitinylation of components of the thick
filament. Interestingly, actin is being degraded in a
specific pattern despite myosin degradation. Thick actin filaments
degrade
earlier than thin actin filaments. Unfortunately, the
question of whether the order of actin filament degradation is
clinically
relevant with regards to the reversibility of the muscle
wasting process remains unanswered. Importantly, the abundant
contractile
protein actin is a target of the UPS in skeletal muscle
both in vitro and in vivo, further supporting the need for new
strategies
to block specifically the activation of this pathway in
muscle wasting conditions [4].

Several investigators studied
mitochondrial dysfunction and its role in muscle wasting. The current
conclusion of these reports
is that mitochondrial dysfunction seems to be the
principal pathway in sarcopenia, i.e. age-related loss of muscle mass [5]. It may be important to differentiate between muscle wasting as part of ‘healthy ageing’ in contrast to muscle loss in chronic
disease, as the involved processes could be fundamentally different [6].
At the meeting, special emphasis was put on possible steps involved in
sarcopenia during the ageing process. Chikwendu
Ibebunjo et al. (Novartis Institutes for Biomedical
Research, Cambridge, Massachusetts, USA) subjected muscle samples to
microarray
and proteomic analysis in a rat sarcopenia model. The
phenotypic, genomic and proteomic features of sarcopenia in these rats
were similar to those of human sarcopenia and suggest
that the animals may provide a suitable model for mechanistic studies
of sarcopenia. Indeed, whilst pathways of mitochondrial
energy metabolism (tricarboxylic acid cycle and oxidative
phosphorylation)
were significantly down-regulated in sarcopenic rats,
genes associated with the neuromuscular junction were up-regulated [7].
Therefore, intervention strategies that counteract these dysregulations
may be beneficial for prevention or treatment of
sarcopenia. In line with this observation is the work of
the group by Siegfried Labeit (Medical Faculty of Mannheim, Mannheim,
Germany) that used MuRF1-knockout mice to analyse the
role of E3 ligase MuRF1 in sarcopenia. Gene inactivation of MuRF1
resulted
in potent protection from muscle atrophy induced by
stimuli such as denervation, hindlimb suspension or injection of tumour
necrosis factor-α (TNFα) [8].
They noted that fast fibre types were preferentially protected.
Furthermore, in line with systemic regulatory effects of
MuRF1 in muscle atrophy, metabolic effects on lipid and
glucose oxidation and circulating amino acid levels could be detected.
Therefore, future studies are warranted to identify
additional pathways that are regulated by MuRF1.

3 Cytokines

The activation of pro-inflammatory
cytokines and a dysbalance between pro- and anti-inflammatory mediators
play important
roles in the pathophysiology of cachexia and muscle
wasting in chronic illness. Cytokines are currently thought of as among
the principal catabolic players in skeletal muscle.

Pedro L. Martinez-Hermandez (Hospital
University La Paz, Madrid, Spain) collected data from 21 cancer patients
and 8 healthy
controls. The authors measured interleukin-15 (IL-15) at
weeks 4 and 8, respectively, because IL-15 might be directly
associated
with changes in body weight [9]. Baseline levels of IL-15 were similar between cancer patients and controls. Whilst cancer patients who gained weight (n = 5) had an increase in IL-15 serum levels, this was not the case in patients who lost weight (n = 13).

Pro-inflammatory cytokines such as IL-6
have likewise seen increasing research efforts over the last several
years. The group
of James A. Carson (University of South Carolina,
Columbia, South Carolina, USA) used the ApcMin/mouse that develops
severe
cachexia and reduced muscle oxidative capacity, to study a
possible role of IL-6 receptor antagonism and exercise in restoring
mitochondrial function in cancer cachexia [10].
They found that whilst loss of muscle oxidative capacity occurs late in
the course of cachexia development, decreased expression
of regulators of mitochondrial biogenesis and dynamics
occur rather early. These data demonstrate clearly that blockade of
IL-6 dependent signalling can reverse deteriorated
mitochondrial function in this model of severe cachexia.

Andrea Bonetto (Thomas Jefferson
University, Philadelphia, USA) and his group concentrated their research
on the signalling
of IL-6 and of signal transducer and activators of
transcription 3 (STAT3), which is known to induce muscle wasting in
cancer
cachexia [11]. Since the
suppressor of cytokine signalling 3 (SOCS3) is one of the inhibitors of
the IL-6/STAT3 pathway, SOCS3 over-expression
in C2C12 myotubes by means of in vitro adenoviral
infection was used to counteract this pathway. Indeed, C2C12 myotubes
that
over-express SOCS3 proved resistant to IL-6-induced
wasting. Furthermore, transgenic mice (MLC-SOCS3 C57BL/6) served to
isolate
the effects of SOCS3 on skeletal muscle. Interestingly,
muscle weight was increased in female MLC-SOCS3 transgenic animals
only, suggesting a gender-specific function of SOCS3. In
cancer cells, over-active receptor and non-receptor-bound tyrosine
kinases cause persistent STAT3 phosphorylation (p-STAT3)
and hence, STAT3 activation. Mice bearing the colon-26 adenocarcinoma
(C26) exhibit severe muscle wasting that was associated
with increased levels of p-STAT3. The researchers demonstrated elegantly
that localized over-expression of SOCS3 in the tibialis
muscle inhibits STAT3 activation and thus prevents muscle wasting
in C26 mice. Therefore, SOCS3 seems to be a possible
therapeutic target in cachexia induced by tumours with high IL-6/STAT3
signalling [11].

4 Therapeutic interventions targeting deleterious cytokine signalling

A very impressive example of the clinical
importance of deleterious IL-6 signalling in cancer cachexia was
reported by Miho
Murakami and colleagues (Wakayama Medical University,
Ibaraki, Japan). They used tocilizumab (TCZ), a humanized anti-human
IL-6 receptor antibody, in a patient with advanced
malignant mesothelioma [12]. In their
case report, a 76-year old man with recurrent malignant mesothelioma of
the pleura was treated with tocilizumab
at a dose of 8 mg/kg of body weight every other
week. The tumour had been refractory to standard polychemotherapy and
radiation,
and therefore treatment with tocilizumab was commenced.
The drug lowered subfebrile temperatures and normalized IL-6-dependent
plasma markers of cachexia like C-reactive protein,
vascular endothelial growth factor and ghrelin within 2 weeks.
Whilst
tocilizumab increased prealbumin and retinol binding
protein, leptin levels were not changed. The general status of the
patient
was considerably improved as documented by the Eastern
Cooperative Oncology Group performance status (ECOG PS scale). Since
the patient suffered from radiation pneumonitis caused by
previous treatment, tocilizumab had eventually been discontinued.
Unfortunately, tumour growth increased rapidly thereafter
[12]. Thus, further studies are warranted to elucidate whether blockade of IL-6 action in patients with malignant mesothelioma
can improve outcomes and possibly the course of cancer cachexia.

Vasilis Paspaliaris (Itis Pharmaceuticals
Pty Ltd., Melbourne, Australia) presented data from a phase I/II study
of IP-1510,
a novel interleukin-1 (IL-1) receptor antagonist that was
used in 26 patients with cachexia caused by advanced gynaecological
cancer. They measured stabilization and increases in body
weight in 17 patients who were treated with IP-1510 at a dose of
1 mg twice daily over 28 days [13].
IP-1510 was well tolerated and safe in patients with advanced cancer.
The interpretation of the current data is limited
because the study was neither randomised nor blinded or
controlled. Large-scale clinical studies are needed to prove whether
neutralization of deleterious cytokines or direct
receptor antagonism is an effective therapeutic approach to improve
patient
outcomes or to reverse muscle loss in cachexia.

5 Therapeutic approaches to counteract immobilisation-induced atrophy

Different studies concentrated on the
mechanisms of muscle wasting induced by immobilisation. A study by L.
Larsson (Department
of Clinical Neurophysiology, Uppsala, Sweden) aimed to
improve the understanding of mechanisms underlying the muscle
type-specific
differences in critically ill patients with acute
quadriplegic myopathy. Using a porcine model, they found an
up-regulation
of UPS activity after 5 days of immobilisation. In
addition, up-regulation of heat shock molecular chaperones as well as a
concomitant down-regulation of sarcomeric proteins such
as MURF2 and growth factors like insulin-like growth factor-1 (IGF-1)
or IGF-2 were reported [14].

Several elegant studies either derived
from animal data or from cell lines were presented with regards to the
role of microRNAs
(miRNA) in skeletal muscle wasting during cancer
cachexia. Each miRNA has multiple target genes (200–500) and thus
represents
an attractive tool to study putative genes involved in
the development of cancer cachexia. Nathan A. Stephen (University of
Edinburgh, Edinburgh, UK) showed that cancer cachexia is
associated with significantly increased expression of skeletal muscle
miR-29b, miR-143, miR-100, miR-768-3p and miR-193b
whereas miR-208a is decreased. Quantitative polymerase chain reaction
(PCR)
validation of these differentially expressed miRNAs
indicated that only miR-29b correlated with weight loss (r = 0.5, p = 0.03) [15, 16]. Therefore, selected miRNAs may represent potential biomarkers for early detection of muscle wasting and targets for future
interventions.

Another approach involves expression
patterns of mRNA in skeletal muscle. Christopher M. Adams (University of
Iowa, Iowa City,
Iowa, USA) showed mRNA expression signatures of human
skeletal muscle atrophy to identify a small molecular inhibitor of
muscle
atrophy. Ursolic acid, a naturally triterpene acid
present in the fatty layer of several fruits and herbs, was used to
demonstrate
reduction in muscle athrophy. For this purpose, the
authors used three distinct mouse models: fasting, denervation and
immobilisation.
They identified 63 mRNAs in human and mouse muscle that
are regulated by fasting in human and mouse muscle, and 29 mRNAs were
regulated by both fasting and spinal cord injury (i.e.
denervation). Their data showed that ursolic acid reduced atrophy and
stimulated hypertrophy by enhancing skeletal muscle
insulin/IGF-1 signalling and inhibiting atrophy-associated mRNA
expression.
Thus, the authors concluded that ursolic acid and the
research on mRNAs may help to prevent and treat muscle atrophy [17].

Jan Vrijbloed (Neurotune AG, Schlieren,
Switzerland) presented a new animal model of sarcopenia, the so-called
SARCO mouse,
a transgenic mouse over-expressing the human enzyme
neurotrypsin. This enzyme produces c-terminal agrin fragment from the
peptide agrin, a synaptically located key player during
initial formation and maintenance of neuromuscular junctions. Levels
of c-terminal agrin fragment are significantly elevated
in a large number of patients with sarcopenia. The SARCO mouse may
therefore offer novel starting points for research of
pathogenic mechanisms in sarcopenia that could be used for
pharmaceutical
treatments [18].

6 MT-102, an anabolic/catabolic transforming agent

Another intriguing approach to the
treatment of muscle wasting was presented by Mareike Pötsch (Charité
Medical School, Berlin,
Germany) who showed data from an animal model using the
new anabolic/catabolic transforming agent MT-102 in order to reverse
muscle wasting in cancer cachexia in the rat. Rats that
were inoculated intra-peritoneally with 108 Yoshida AH-130 hepatoma
cells and were then treated with MT-102 at a dose of
3.0 mg/kg/day resulted in a gain of lean mass and body weight. On
day 11,
left ventricular ejection fraction (p < 0.01), fractional shortening (p < 0.05), and stroke volume (p < 0.01) were significantly improved by MT-102 [19].
The ACT-ONE trial, a multicentre, randomised, double-blind,
placebo-controlled, dose-finding study of the anabolic/catabolic
transforming agent MT-102 that has recently commenced
recruitment of subjects with cachexia related to stages III and IV
non-small
cell lung cancer or colorectal cancer needs to confirm
whether these animal data can be translated into humans [20].

7 Clinical studies/growth hormone

Several factors have complicated the
development of effective therapies for cachexia particularly the many
different pathways
involved in the development of the disease. The focus of
many therapies includes nutritional interventions. There has been
little consensus on the primary end point for clinical
trials, which has hampered assessment of the efficacy of treatments.
Results of randomised, double-blind, placebo-controlled
adequately powered clinical trials in the field of cachexia are eagerly
awaited.

A promising approach was presented by John
Friend (Helsinn Therapetuics Inc, Bridgewater, New Jersey, USA) who
provided insight
into the design of a randomised, double-blind,
placebo-controlled, multicenter phase III study to evaluate the safety
and
efficacy of anamorelin in patients with non-small-cell
lung carcinoma (NSCLC). Anamorelin is an orally active ghrelin receptor
agonist. Ghrelin is a physiological ligand of the growth
hormone secretagogue receptor that stimulates food intake. Anamorelin
as a potential ghrelin-analog seems to be a therapeutic
agent with possible effects during all stages of cachexia. Study results
are estimated to be presented at the 7th Cachexia
Conference in Kobe in December 2013.

Plasma ghrelin levels are elevated in cachectic patients with chronic heart failure possibly through a compensatory mechanism
to a catabolic/anabolic imbalance [21, 22].
Exogenously administered ghrelin has been shown to improve left
ventricular dysfunction and to attenuate the development
of cardiac cachexia in rats with heart failure. Thus,
supplementation of ghrelin could be a therapeutic approach in the
treatment
of chronic heart failure [23, 24].
A naturally occurring splice variant of ghrelin (Dln-101) with 57%
homology to ghrelin was presented. Liat Mintz (DiaLean
Ltd., East Brunswick, New Jersey, USA) conducted
extensive pre-clinical studies showing that Dln101 acts similarly to
ghrelin
in increasing food intake, promoting weight gain and
increasing growth hormone release. In addition, Dln-101 suppresses
inflammation
and has beneficial effects on the metabolic profile in
terms in reducing cholesterol and glucose levels. Thus, Dln-101 has
received approval to start phase I clinical trials [25].

8 Erythropoietin

Yulia Elkina (Charite Medical School,
Berlin, Germany) and colleagues showed the tissue-protective effect of
the non-hematopoietic
erythropoietin analogues ARA284 and ARA286 in the
treatment of cancer cachexia in a rat cancer cachexia model. ARA284 and
ARA286 in high concentrations (5,000 units/kg/day)
were shown to be effective in reducing tissue wasting in rat cancer
cachexia
model. These compounds should be seen as prospective
drugs for human cancer cachexia [26].

9 Selective androgen receptor modulators

Selective androgen receptor modulators
(SARMs) belong to a relatively new class of therapeutics that possesses
anabolic properties.
SARMs are currently in the early stages of development [27]. A double-blind, placebo-controlled phase II clinical trial with GTx-024 (enobosarm) in 120 healthy elderly men and women
showed a dose-dependent (3 mg of enobosarm) improvement in total lean body mass and physical function [28]. Shontelle Dodson (GTx Inc., Memphis, USA) and colleagues analysed enobosarm that was used in the treatment of 159 patients
with NSCLC with muscle wasting [29].
The group that received enobosarm showed positive effects on quality of
life. Additionally, they found a benefit with regards
to stair climbing power, a readily available measure of
exercise capacity, frequently used by geriatricians. Moreover, stair
climbing power benefit (defined as 10% improvement)
correlated with quality of life (measured with functional assessment of
cancer therapy). According to that, enobosarm had an
impact on physical function, quality of life, and it had the ability
to overcome the negative prognostic effect of >8%
weight loss on overall survival [30].
Eric Vajda (Ligand Pharmaceuticals, La jolla, California, USA) presented
data from another novel SARM termed LGD-4033 that
was used in healthy young men. In this randomised,
placebo-controlled study, the dose of 0.1, 0.3 or 1 mg LGD-4033
daily was
studied over a time of 21 days. This treatment phase
was followed by a 5-week observation period. The study showed a
dose-dependent
increase in lean body mass (about 1.2 kg) in
patients who were treated with 1 mg LGD-4033 [31].
The investigators are planning a phase II study to evaluate LGD-4033 in
conditions such as muscle wasting associated with
cancer and acute illness. The mechanism by which
steroidal and non-steroidal SARMs produce selective tissue-specific
anabolic
responses has not been fully elucidated. However, further
studies are necessary to evaluate the mechanism and long-term effect
of SARMs in patients with cachexia.

10 Conclusions

Several pathways have been shown to play
important roles in muscle wasting in the muscle wasting process: actin
as a target
of UPS, MuRF-regulated pathways or IL-15 that may be
directly associated with loss of body weight. Potential biomarkers for
cachexia are currently under investigation. There is
further need for attractive biomarkers as therapeutic targets in the
context of cachexia. The outcome criteria of drug studies
in cancer cachexia need to focus not only on mortality, but also
on symptoms and quality of life rather than simply on
nutritional end-points, since the survival of cachectic cancer patients
is usually limited to weeks or months due to the
incurable nature of the underlying malignancy. In summary, cachexia and
sarcopenia
need more attention in clinical work and research and
prospective clinical randomised, double blinded, placebo-controlled
studies are needed. Prospective large-scale studies are
warranted, such as the Studies Investigating Co-morbidities Aggravating
Heart Failure (SICA-HF), a multicenter pathophysiological
observational study which is particular looking at cachexia and
obesity prevalence in chronic heart failure [32]. The results thereof will be presented in Kobe in 2013.

Acknowledgement The authors of this manuscript certify that they comply with the ethical guidelines for authorship and puplishing in the Journal
of Cachexia, Sarcopenia and Muscle [33].

Part of this work has received funding from the European Union Seventh Framework Programme [FP7/2007-2013] under grant agreement
n° 241558 (SICA-HF).

Conflict of interest

The authors declare that they have no conflict of interest.

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